Novel 21-acyloxy-3alpha-hydroxy-5alpha-pregnone-11,20-diones

ABSTRACT

THIS INVENTION IS CONCERNED WITH IMPROVEMENTS IN OR RELATING TO PHARMACEUTICAL PREPARATIONS HAVING ANAESTHETIC ACTIVITY, AND MORE PARTICULARLY AQUEOUS PREPARATIONS SUITABLE FOR INTRAVENOUS INJECTION.

United States Patent Int. Cl. C07C 169/32 US. Cl. 260-397.45 7 ClaimsABSTRACT OF THE DISCLOSURE This invention is concerned with improvementsin or relating to pharmaceutical preparations having anaestheticactivity, and more particularly aqueous preparations suitable forintravenous injection.

This application is a division of co-pending application Ser. No.47,163, filed June 17, 1970.

It has long been known that a number of steroids give rise to profounddepression of the central nervous system and act pharmacodynamically asanaesthetics or hypnotics. Such compounds have been the subject ofconsiderable study in an atempt to find anaesthetics to replace suchsubstances as thiopental sodium normally used but well known to beaccompanied by some degree of hazard or disadvantage. The literatureshows that very many steroid compounds have been studied in this regard.Reviews and discussions of some of the work carried out are to be found,for example, in Methods in Hormone Research (Edited by Ralph I. Dorfman,vol. III, Part A, Academic Press London and New York 1964, pages 415-475); H. Witzel, Z Vitamin, Hormon-Fermentforsch 1959, 10, 46-74; H.Selye, Endocrinology, 1942, 30, 437- 453; S. K. Figdor et al., J.Pharmacol. Exptl. Therap., 1957, 119 299-309 and Atkinson et al., J.Med. Chem. 1965, 8, 426-432.

A thorough review of the literature indicates that anaesthetic steroidsgenerally possess poor activity and/ or long induction periods. Withsuch compounds a variety of undesired side effects such as paraesthesiaand thrombophlebitis and vein damage have been noted. Many steriodcompounds having anaesthetic action are also of poor solu-bility andthus much research has hitherto been directed to the introduction ofsolubilising groups into such steroids, e.g. by the formation of partialesters with dior polybasic acids; such work has hitherto not resulted inthe discovery of a satisfactory anaesthetic steroid compound.Anaesthetic steroids are generally relatively simple pregnanederivatives, often hydroxylated in the 3 position, the general trendhaving been in the latter case to study 3,8-hydroxy compounds inpreference to 30- hydroxy compounds.

As a result of prolonged study of numerous steroids exhibitinganaesthetic activity we have found that 304- hydroxy 50c pregnane 11,dione (hereinafter called steroid I) has quite remarkable properties asan anaesthetic. This substance has but poor solubility in Water andalthough referred to in the above cited literature as having anaestheticproperties, has apparently been rejected by reason of its poorsolubility and the difficulty to be anticipated in its use byintravenous administration arising therefrom.

We have found however that steroid I can be brought into aqueoussolution with the aid of certain non-ionic surface active agents toyield solutions of a sufiicient con- 3,763,195 Patented Oct. 2, 1973 icecentration for injection, the resulting solutions when injected showingexcellent anaesthetic properties.

Thus the aqueous solutions of steroid I according to the inventioninduce anaesthesia and possess short induction periods, the anaestheticaction at suitable doses being indeed instantaneous; the solutions arethus excellent anaesthetics for inducing anaesthesia which is to bemaintained e.g. by an inhalation anaesthetic such as ether, halothane,nitrous oxide, trichloroethylene etc. The solutions are however capableof maintaining anaesthesia and analgesia to a suflicient degree toenable various surgical operations to be conducted without the aid of aninhalation anaesthetic, the required degree of anaesthesia beingmaintained if necessary by repeated administration (or even continuousadministration). Recovery from anaesthesia (where this is induced onlyby the solutions of this invention) is excellent, the patient exhibitinga feeling of well-being in distinction to the unpleasant after effectsgenerally associated with conventional anaesthetics. Moreover,anaesthetic solutions in accordance with the invention in general giverise to none of the undesired side-effects hitherto associated withsteroidal anaesthetics.

According to the invention therefore We provide pharmaceuticalcompositions suitable for use by parenteral administration, comprisingan aqueous solution of at least 1 mg./ml. of3a-hydroxy-5a-pregnane-11,20-dione and at least 1% by weight of aparenterally acceptable nonionic surface active agent having an HLBvalue of from 9-18.

The non-ionic surface active agents used for the purpose of thisinvention are generally those of the water soluble type, preferablyhaving an HLB value of at least about 12, advantageously at least about13. Preferably the HLB value of the surface active agent is not greaterthan about 15. The surface active agent must naturally be one which isphysiologically compatible, i.e. of itself give rise to nophysiologically unacceptable side effects in the dosages employed in theintended species to be treated (man or animal). Surface active agentsfor use in accordance with the invention are for example to be foundamong the following non-ionic surfactants and classes ofsurfactants:P0lyoxyetl1ylated derivatives of fatty (C12- C20) glycerideoils, e.g. castor oil, containing from 35 to 45 oxyethylene groups, permole of fatty oil. Polyoxyethylene ethers (containing from 10 to 30polyoxyethylene groups) of long chain alcohols (containing for examplefrom 12-18 carbon atoms).

Polyoxyethylene-polyoxypropylene ethers containing from 15 to 35 andfrom 15 to 30 oxyethylene and oxyethylene and oxypropylene groupsrespectively. Polyoxyethylene ethers (containing from 6 to 12oxyethylene groups) of alkyl phenols the alkyl groups of whichpreferably contain 6-10 carbon atoms.

Polyoxyethylated (containing from 15 to 30 oxyethylene groups) fattyacid (e.g. Cl2-18) esters of sugar alcohol anhydrides e.g. sorbitan ormannitan. Long-chain (e.g. Cl0-l6) alkanoyl monoand di-alkanolamides(the alkanol portions of which for example contain 1-5 C atoms) forexample lauroyl monoand di-ethanolamides. Polyethylene glycol esters(containing from 6 to 40 ethylene oxide units) of long chain fatty acids(containing for example 12-18 C atoms) e.g. polyethyleneglycolmono-oleate (containing for example 8 ethylene oxide units).

Examples of non-ionic surface active agents, of the foregoing types,useful in accordance with the invention include:

Cremophor EL, a polyoxyethylated castor oil containing about 40ethyleneoxide units per triglyceride unit;

Tween 80, polyoxyethylene sorbitan monooleate containing about 20ethylene oxide units;

Tween 60, polyoxyethylene sorbitan monostearate containing about 20ethylene oxide units; and

Tween 40, polyoxyethylene sorbitan monopalmitate containing about 20ethyleneoxide units.

The expression solutions is used herein to denote liquids which have theappearance of true solutions and are thus optically clear and capable ofpassage, for example, through a micro-porous filter, irrespective ofwhether such solution are true solutions in the classical chemical senseand irrespective of whether they are stable or metastable. Thus it maybe that the steroid is associated with micelles. The solutions of thisinvention, irrespective of their precise physical nature, behave as truesolutions for the practical purpose of intravenous injection.

We have further found that the total amount of anaesthetic steroid whichmay be dissolved in accordance with the invention may be substantiallyincreased by the presence of a steroid of general formula CH2OH whereinR is an alkanoyl group having a straight or branched chain (containingfor example 2-4 carbon atoms which may, if desired be substituted, forexample by a carboxyl group) or an unsubstituted or substituted aroyl oraralkanoyl group. Such steroids of Formula II thus act as solubilitypromoters for the steroid I and are capable of substantially increasingthe amount of the latter which may be dissolved in the compositions ofthis invention. Moreover steroids of Formula II, preferred members ofwhich are those in which R is an acetyl, propionyl, isobutyryl,hemisuccinoyl or benzoyl group, themselves possess anaesthetic activityalthough of a generally lower order than steroid I. Solutions containingsteroid I together with a steroid of Formula 11 may thus be preparedhaving a substantially greater anaesthetic potency than a solution ofsteroid I alone, due to the increased amount of steroid I which can bedissolved and the anaesthetic action of the steroid of Formula II perse.

Compounds of Formula II above are new compounds, and constitute afurther feature of the invention.

The proportion of surface active agent to be used in the compositions ofthis invention depends upon its nature and upon the concentration ofsteroid desired in the final composition.

In preferred compositions according to the invention the proportion ofsurfactant is preferably at least 5% by weight and advantageously aboveby weight. A very convenient proportion of surfactant has been found tobe by weight but 30% and up to 50% may be used. The proportions ofsurfactant are expressed by weight in relation to the total volume ofthe composition in metric units.

The compositions according to this invention may contain up to 40 oreven 50% by Weight of the total steroid of a compound of Formula II; aslittle as 5% of the total steroid present of a compound of Formula IIprovides useful solubility promotion. For practical purposes it ispreferred that of the total steroid at least 10% by weight, andpreferably not more than 30% by weight of the total steroid consists ofa compound of Formula II. Very satisfactory results have been achievedwith a mixture of steroid I with a steroid of Formula II in which theproportion of the steroid of Formula II is about As will be clear, theproportion of steroid I in the aqueous solution according to theinvention depends upon the nature and amount of surface active agentused and also upon the amount of steroid of Formula II present (ifused). The composition will contain at least 1 mg./ml. of steroid I andsolutions can be made containing for example 2 and 4 mg./ml.; using asteroid of Formula II as solubiliser the amounts of total steroid can beincreased up to 50 mg./ml. although less than 30 mg./ml. total steroidand generally less than 20 mg./ml. will usually be satis factory. Acomposition comprising 9 mg./ml. of steroid I to 3 mg. /ml. of acompound of Formula II in which R is acetyl has been found to be verysatisfactory.

In all cases, as stated above the relative proportion of the variouscomponents are adjusted to give a clear solution.

In a preferred method of preparing the solutions according to theinvention the steroid is first dissolved in the selected surfactant forexample, with heating and the resulting solution dissolved in water.Alternatively the steroid may be dissolved in a volatile organic solventadvantageously having a boiling point of less than about C. which ismiscible with the surface active agent such as a volatile loweraliphatic ketone, e.g. acetone or methyl ethyl ketone or a volatilehalogenated hydrocarbon, e.g. chloroform or methylene chloride. Acetoneis particularly preferred for this purpose. The surface active agent isthen added to this solution, the organic solvent removed by evaporation,for example by passing a stream of an inert gas through the solution,e.g. nitrogen and the resulting solution of steroid in surfactant ismixed with water. By these procedures in general it is possible todissolve increased amounts of steroid I as compared with other methods.

The solutions may also be prepared by shaking the steroid with anaqueous solution of the surface active agent.

In all cases simple tests enable one to determine the relativeproportions of surface active agent required.

The anaesthetic solutions according to the invention are generallyadministered by intravenous injection although as is known in theanaesthetic art in certain cases, e.g. with young children intramuscularinjection might 'be preferred.

As is usual in the case of anaesthetics, the quantity of steroid I usedto induce anaesthesia depends upon the weight of the individual to beanaesthetised. For intravenous administration in the average man a doseof from 0.45 to 3.5 mg./ kg. will in general be found to be satisfactoryto induce anaesthesia, the preferred dose being within the range of from0.7 to 2.5 mg./kg. Generally a dose of about 1.35 mg./ kg. is verysatisfactory. The dose will naturally vary to some extent dependent uponthe physical condition of the patient, and the degree and period ofanaesthesia required, all as is well known in the art. It is thuspossible by adjustment of the dose to achieve durations of anaesthesiavarying from about 10 minutes to up to an hour or more. If it is desiredto maintain prolonged anaesthesia, repeated doses of the solutions ofthis invention may be used, such repeated doses being generally eitherof the same order or lower than the original dose. Alternativelycontinuous administration may be undertaken at for example a rate of0.09l.8 mg./kg./min.

Where the anaesthetic solutions are administered intramuscularly,naturally higher doses are generally necessary.

3a-hydroxy-5a-pregnane-l1,20-dione (steroid I) can be prepared in anyconvenient manner for example as described by Camerino et al., Helv.Chim. Acta., 1953, 36, 1945 or by Nagata et al., Helv. Chim. Acta. 1959,42, 1399. In applying the last-mentioned process we have encounteredvarious difficulties at several stages'due to side reactions, which wehave found to be due to epimerisation at position 17. As a result offurther work we have developed a new synthesis of steroid I basedessentially upon the synthesis of Nagata et al. but working essentiallywith intermediates having A -unsaturation. Our new synthesis providesgenerally better yields and affords greater ease of working and purityof product, due essentially to the impossibility of epimerisation atposition 17 in compounds of the A series.

Thus in accordance with our new synthesis 3oc-l1YdIOXY-5a-pregnane-11,20-dione is prepared by hydrogenation of3a-hydroxy-5a-pregn-16-ene-11,20-dione.

The hydrogenation may be conveniently effected in an organic solvent,for example an alkanol or an ether e.g. methanol, ethanol, propanol,diethyl ether or tetrahydrofuran. A catalyst is preferably used toeffect the hydrogenation for example palladised charcoal, Raney nickel,platinum catalysts and the like.

3a-hydroxy-5ot-pregn 16-ene-l1,20-dione may, for example, be prepared bysolvolysis of the 3a-acyloxy group of a3a-acyloxy-5a-pregn-16-ene-l1,20-dione. The acyloxy group at theSet-position may, for example, be a formyloxy, acetoxy, propionyloxy,butyryloxy or benzoyloxy group.

Advantageously, the solvolysis may be effected by hydrolysis, preferablyunder alkaline conditions although both acidic and alkaline hydrolysesmay be used. The hydrolysis may, for example, be effected in thepresence of a solvent such as an alkanol or an ether, e.g. methanol,ethanol, propanol, iso-propanol, butanol, t-butanol, diethyl-ether,dioxan or tetrahydrofuran. The reaction may be effected at anyconvenient temperature, for example, at ambient temperature or anelevated temperature, advantageously at the boiling point of the solventused. Suitable alkalis for the hydrolysis include, for example, alkalimetal hydroxides e.g. sodium or potassium hydroxide.

The 3a-acyloxy-5m-pregn-16-ene-11,20-diones which may be used for thepreparation of the corresponding 30:- hydroxy compound may, for example,be prepared from a corresponding 3fi-hydrocarbonsulphonyloxy-5a-pregn-16-ene-ll,20-dione e.g. 3/3-tosyloxy-5a-pregn-16-ene-11, 20-dione andthis reaction will be seen to be an inversion reaction in which theconfiguration at position-3 is changed from [3 to or.

According to this method, the 3B-sulphonate may be reacted with theappropriate carboxylic acid in solution in a solvent medium to yield thedesired 3a-acyloxy compound. This reaction is preferably effected in thepresence of alkali metal ions it being convenient to use the carboxylicacid in the form of an alkali metal salt thereof e.g. the sodium orpotassium salt. Suitable solvents include for example dimethylsulphoxideand N- alkylamide solvents, e.g. dimethylformamide anddimethylacetamide. Preferably the solvent is water miscible and containswater (for example up to 20%) which general- 1y serves to accelerate thereaction. Advantageously the carboxylic acid is an aliphatic carboxylicacid, lower alkanoic acids being preferred, for example, formic, acetic,propionic or butyric acids or an aromatic carboxylic acid e.g. benzoicacid. Preferred salts are the sodium and potassium salts. Generallyfavourable results are obtained by the use of potassium acetate. Thesolvent used may be the carboxylic acid itself or an anhydride thereof(provided that such acid or anhydride is liquid at the reactiontemperature) alkali metal ions being provided by an alkali metal salte.g. of the carboxylic acid. For example when using aqueous formic acid,alkali metal ions which may be present may, for example, be provided byan alkali metal formate or an alkali metal hydrogen carbonate e.g.potassium formate or potassium hydrogen carbonate. The inversion of theconfiguration of the substituent at the 3-position of the 5a-pregn-16-ene-11,20-dione may be effected at any convenient temperature,advantageously at an elevated temperature, suitable conditions beingfound by preliminary experiment.

An alternative method of converting the above-mentioned 3/3-sulphonatesto the corresponding 3a-hydroxy compounds involves treatment of thesulphonates under alkaline conditions using for example an alkali metalhydroxide, e.g. NaOH, in a polar solvent e.g. ethanol, methanol, dioxanetc.

The inversion of the configuration of the substituent from the3,8-position to the 3a-position may also be effected in the presence ofdimethylformamide, or dimethylacetamide preferably in the presence ofwater e.g. analogous to the method described by Chang and Blickenstaff(J. Amer. Chem. Soc., 1958, 80, 2906), and Bharucha et al. (Can. J. Chem1956, 34, 982). This method when applied, for example, to the tosylateof 3,B-hydroxy- 5a-pregn-16-ene-11,20-dione produced the corresponding3a-formate together with some of the corresponding free hydroxycompound. Dimethyl acetamide may also be used, preferably under aqueousconditions, to form the 3oc-8C6tOXy compound by inversion.

3 B-hydrocarbon sulphonyloxy-Sa-pregn-l6-ene 11,20- diones may, forexample, be prepared according to conventional methods. Thus the Sd-hydroxy compound may be reacted with a hydrocarbon sulphonyl halide,for example p-toluene sulphonyl chloride, preferably in the presence ofa tertiary base e.g. pyridine, collidine, N- methyl morpholine etc., thetertiary base, if desired, also acting as solvent. The sulphonylationreaction may, for example, be effected at a temperature of from 50 to 50C., preferably 0 to 25 C.

In carrying out the process according to the invention for thepreparation of 3a-hydroxy-5a-pregnane-11,20- dione, good overall yieldsfrom 3fi-hydroxy-5u-pregn-16- ene-1l,20-dione may be obtained by theomission of purification of at least some of the intermediates. Thisprocess also has the advantage of comparative ease of purification of3a-hydroxy-5ot-pregnane-l1,20-dione from the corresponding 3-desoxycompound formed as a by-product in the process from the corresponding2,16-diene formed by elimination of the hydrocarbon sulphonyloxy groupin the inversion step. Good overall yields from3B-hydroxy-5apregnane-11,20-dione of high purity, may be achieved usingpotassium acetate in aqueous dimethylformamide in the inversion step.Improved results may be obtained by partial purification of the reactantused in the hydrogenation step.

3a-hydrOXy-Sa-pregn-I6-ene-11,2O-dione may also be prepared by hydrationof 5ot-pregna-2,16-diene-11,20- dione which may be formed by theelimination of the hydrocarbon sulphonyloxy group from the correspondingZip-hydrocarbon sulphonyloxy compound. The hydration may, for example,be effected to the reaction with a mercury compound, preferably underacid conditions to form an oxymercurial intermediate which is thendecomposed by reduction. For example, the oxymercuration may be carriedout using mercuric acetate, mercuric sulphate or mercuric oxide in asolvent such as dioxan or tetrahydrofuran (preferably aqueous)advantageously in the presence of a strong acid e.g. perchloric acid,nitric acid, or sulphuric acid. Decomposition of the oxymercurialintermediate may for example be effected by reduction with an alkalimetal borohydride e.g. sodium or potassium borohydride, preferably inthe presence of aqueous alkali e.g. aqueous caustic soda.

3fl-hydroxy-5ot-pregn-16-ene-11,20-dione may be prepared from3B-acetoxy-5u-pregn-16-ene 11,20 dione (Chamberlin et al., J. Amer.Chem. Soc., 1951, 73, 2396). Hydrolysis of the 3B-acetoxy compound maybe effected by conventional methods, for example under alkalineconditions, e.g. in solution in acetone, an alkanol or ether such asmethanol, ethanol, propanol, t-butanol, diethyl ether, dioxan ortetrahydrofuran. Suitable alkalis may, for example, be alkali metalhydroxides e.g. potassium hydroxide.

Steroids of General Formula II above are new compounds and constituteanother feature of the invergion. The preferred compounds of Formula IIfor use as solubilising agents in pharmaceutical preparations are thosein which the 2l-acyloxy group is a 21-acetoxy, 21-propionyloxy,2l-isobutryloxy or 2l-benzoyloxy group.

The 2l-acyloxy compounds of Formula II above may be prepared by anyconvenient method. We have found however that such compounds areadvantageously prepared by reaction of a compound of formula COCHa H(III) (in which R represents a hydroxy or protected hydroxy group e.g. anitrate, trimethylsilyloxy or trichloroethoxycarbonyloxy group) with alead tetraacylate preferably in the presence of a Lewis acid, and, whereR represents a protected hydroxy group, subsequent conversion of saidgroup to a hydroxy group. Protection of the 3a-hydroxy group of thecompound of Formula III however, is not essential before theacyloxylation reaction.

The lead tetra-acylate used may be for example lead tetraacetate. TheLewis acid may for example be boron trifiuoride, conveniently used asits etherate. We have found, for example that the presence of borontrifiuoride improves the rate of reaction and can enable lowertemperatures of reaction to be employed, thus, in many cases, thereaction proceeds satisfactorily at ambient temperature, i.e. in theabsence of applied heat.

The yield obtained by this process is frequently better than thatobtained at an elevated temperature in the absence of boron trifiuorideand the fact that the reaction rate is faster and thus lowertemperatures can often be used means that this process possesseseconomic advantages on the large scale. The ability to operate at lowertemperatures also means that there may be less likelihood of undesiredside reactions taking place.

The acyloxylation of compounds of the Formula III may be carried out ina solvent medium comprising a mixture of a hydrocarbon solvent and analcohol. Suitable hydrocarbon solvents are, for example, benzene ortoluene and the alcohol may, for example, be methanol. Advantageouslythe solvent comprises a mixture of benzene and methanol in the ratio of19:1.

The acylroxylation is especially suitable for acetoxylation using leadtetraacetate but other lead acylates, e.g. lead tetrapropionate may, ofcourse, be used with the formation of the corresponding 21-propionyloxycompound.

The 3ot-trimethylsilyloxy compounds of Formula III (R representing atrimethylsilyloxy group) may, for example, be prepared by the reactionof the parent 3tx-hydroxy compound of Formula III (R' representing ahydroxy group) with a trimethylsilyl halide e.g. trimethylchlorosilaneor hexamethyl disilazane in the presence of a tertiary base e.g.pyridine and, if desired, in the presence of a solvent e.g. ahalogenated hydrocarbon such as methylene chloride or tetrahydrofuran.The reaction may conveniently be effected at room temperature or ifdesired, at lower temperatures e.g. C. The trimethylsilyloxy pro tectinggroup is generally automatically removed during the acyloxylation.

The 3a-trichloroethoxycarbonyloxy compound of Formula III may beprepared by reaction of the parent 30thydroxy compound with analkylchloroformate, e.g. trichloroethyl chloroformate, preferably in thepresence of an acid binding agent e.g. a tertiary amine such aspyridine, conveniently in a solvent such as dioxan or tetrahydrofuran.The trichloroethoxy carbonyl protective group may subsequently beremoved by reduction, for example, using a metal acid system such aszinc and acetic acid or catalytic hydrogenation using for example,palladium on charcoal as catalyst.

Other methods for the preparation of the 2l-acyloxy comp unds of FormulaII above may also be used. Thus a 21-acyloxy-5a-pregnane-3,11,20-trionemay be reduced, for example, using an enzymatic method such as reductionwith brewers yeast (Saccharomyces cerevisiae). This method is convenientfor the preparation of 21-alkanoyloxy compounds such as the 21-acetoxycompound.

Compounds of Formula II may also be prepared via the corresponding2l-chloro, 2l-bromo or 21-iodo compounds. We prefer to proceed via the21-bromo intermediate which may be prepared from 5u-pregnan-3a-ol-11,20-dione. The bromination is effected for example using molecularbromine, in a solvent such as methanol or ethanol advantageously at atemperature of from -10 to +30 C. The reaction is preferably conductedin the presence of a catalyst such as acetyl chloride or hydrogenbromide in acetic acid. The 21-halogeno compound may then be convertedinto the desired 21-acyloxy compound by reaction with the salt ofcorresponding carboxylic acid, such as an alkali metal salt e.g. thepotassium salt or a tertiary amine salt conveniently N-methylmorpholineor N-ethylpiperadine or a trialkylammonium salt e.g. thetriethylammonium salt. The reaction is preferably carried out in asolvent for example acetone or methanol, advantageously under anhydrousconditions. An alternative method, particularly convenient for thepreparation of compounds of the Formula II in which R is other than anacetyl group, comprises acylating a compound of formula of R (wherein Rrepresents a protected hydroxy group) and deprotecting the 3a-hydroxygroup of the Zl-acyloxy derivative of the compound of Formula IVproduced. The compound of Formula IV is conveniently prepared by thedeacylation of a 2l-acyloxy derivative of a compound of Formula IV, forexample a 21-acetoxy derivative thereof, and provides a method forconverting one compound of Formula II for example in which R is acetylto another compound of Formula II in which R is a different acyl group.The 21-acetyl compound of Formula II may thus, for example, be convertedinto another 2l-acyl compound of Formula II by firstly protecting the3:1- hydroxy group with a protecting substituent which may be removedunder acidic, reductive or other conditions but is stable under alkalineconditions, such as a Set-ether substituent e.g. a tetrahydropyranyl ortriphenylmethyl substituent or preferably a Zia-nitrate ester,hydrolysing the 3a-protected compound to yield the corresponding21-hydroxy compound, under basic conditions, preferably in the presenceof potassium or sodium hydrogen carbonate, conveniently in the presenceof a solvent e.g. methanol, ethanol or tetrahydrofuran, reesterifyingthe resultant product and removing the But-protecting substitutent.

The reesterification is preferably effected using the an hydride orchloride of the desired acid preferably in the presence of a teriaryamine (e.g. pyridine, collidine, or dimethylaniline) which may alsoserve as solvent for the reaction.

The protecting group at position 3 may be removed in conventionalmanner, conditions may be chosen which will not effect the rest of themolecule. Thus for example when the 3a-hydroxy group of the compound ofFormula II is protected by the formation of a nitrate ester, the nitrategroup may be removed by acid hydrolysis of the compound for exampleusing aqueous mineral acid, or by reduction, using, for example zinc andacetic acid or by COCHzOH 9 catalytic hydrogenation, using, for example,palladium on charcoal as catalyst.

The following examples are given by way of illustration only,alltemperatures being in degrees centigrade.

The following examples are given by way of illustration only, alltemperatures being in degrees Centigrade.

EXAMPLE 1 3 B-hydroxy-ot-pregn-16-ene-1 1,20-dione A solution of3fi-acetoxy-5a-pregn-l6-ene-l1,20-dione (Chamberlin et al., J. Amer.Chem. Soc., 1951, 73 2396) (25.7 g.) in dioxan (Analar, 500 ml.) wastreated with potassium hydroxide g.) and water 250 ml. and the mixtureallowed to stand at room temperature for 1 hour. After a further 1 hourat 40 the mixture was diluted with water and the product filtered off.The crude material Was dissolved in chloroform and filtered through acolumn of grade III neutral alumina (ca. 100 g.). The material obtainedwas crystallised from acetone-petroleum to give pure3,8-hydroxy-5a-pregn 16 ene 11,20- dione (17.65 g., 77.5%) as smallplates, M.P. 217.5", [0L]D+82-9 (C., CHC1 1.1), k 234 nm. (6 10,00).(Found: C, 75.9; H, 9.3. C H O requires C, 76.3; H, 9.2%).

EXAMPLE 2 Bfi-toluene-p-sulphonyloxy-5ot-pregn-16-ene-11,20-dione Asolution of 3fi-hydroxy-5u-pregn-16-ene-11,20-dione (39.6 g.) in drypyridine (165 ml.) was treated with toluene'p-sulphonyl chloride (43.9g.) to give the toluene sulphonate (56.7 g.) M.P. l47-151. A portion(10.7 g.) of this material was crystallised from ethyl acetatepetroleumto give the pure toluenesulphonate (9.2 g.) as plates M.P. 154-155, [a]+42.8 (C., CHCl 1.2), Amax 226 nm. (6 20,780). Found: C, 69.6; H, 7.4;S, 6.5. C2gH3 05S requires C, 69.45; H, 7.5; S, 6.6%).

EXAMPLE 3 3a-hydroxy-5a-pregn-16-ene-11,20-dione A solution of3,B-toluene-p-sulphonyloxy-5a-pregn-l6- ene-ll, -dione (19.1 g.) inN,N-dimethylformamide (160 ml.) and water (16.0 ml.) was treated withpotassium acetate (29.2 g.) and the mixture heated at 115 (completesolution on heating) for 2 /2 hours. The solvents were removed in vacuoand the residue partitioned between chloroform and water. The chloroformextract was washed with water, dried and evaporated. The residue wastaken up in methanol (500 ml.) and the solution flushed with nitrogen.Potassium hydroxide (17 g.) in water (70 ml.) was added and the solutionrefluxed for 1 hour. Glacial acetic acid was added to bring the pH toabout 6 and most of the methanol evaporated in vacuo, Dilution withwater gave a gummy precipitate which was extracted into chloroform togive the crude product. This material was extracted with ether and theresidue boiled with benzene. The insoluble material was crystallisedfrom chloroform-petroleum to give 3ot-hydroxy-5a-pregn-16-ene-1l,20-dione (3.28 g.) as large prisms M.P. 243- 244", [a] +86.5(C.,CHCl 0.8), Amax' 233 nm. (5 9,530). (Found: C, 76.6; H, 9.2. C H Orequires C, 76.3; H, 9.15%).

EXAMPLE 4 3ohydroxy-5ot-pregnane-11,20-dione From3a-hydroxy-5ot-pregn-l6-ene 11,20 dione.A solution of3ot-hydroxy-5u-pregn 16 ene 11,20 dione (200 mg.) in freshly distilledtetrahydrofuran (8 ml.) with 5% palladium on carbon (100 mg.) washydrogenated till hydrogen uptake ceased. The mixture was filteredthrough a pad of kieselguhr and the tetrahydrofuran removed in vacuo togive 3u-hydroxy-5a-pregnane- 11,20-dione (196 mg.) M.P. 171-172", [a]+112.5 (C., CHCl; 1.0).

10 EXAMPLE 5 3a-hydroxy-5ot-pregnaue-l1,20-dione A solution of 38-acetoxy-5a-pregn-16-ene-11,20-dione (50 g., 0.134 mole) in dioxan(1,050 ml.) was flushed with nitrogen and treated with potassiumhydroxide (20 g., 0.356 mole) then nitrogen flushed water (535 ml.). Thesolution was stored at 40 for 1% hours then at room temperature for 1hour. Glacial acetic acid was added (to pH ca. 7), the organic solventremoved in vacuo until crystallisation started and the resulting mixturediluted with water (ca. 1,500 ml.). The precipitated solid was filteredoff, washed with water (until washings were neutral) and dried overphosphorus pentoxide in vacuo. This gave the alcohol (39.9 g., theory)as an off-white crystalline solid M.P. 216-218", [d] +82.9 (c. 1.0, CHCl'ymax, 234 nm. (6 9,210).

An ice-cold solution of toluene-p-sulphonyl chloride (96 g., 0.505 mole)in dry pyridine (180 ml.) was added to an ice-cold solution of the abovealcohol (86 g., 0.26 mole) in dry pyridine (180 ml.). The dark orangesolution was allowed to warm to room temperature and kept at thattemperature for 16 hours. The mixture was cooled to 0 and sufficientwater (ca. 20 ml.) added to dissolve the precipitated pyridinehydrochloride. The clear solution was kept at room temperature for 2hours then diluted with 2 N-hydrochloric acid (2.5 1.). The precipitatedcrystalline solid was filtered off, washed with 2 N-hydrochloric acid(500 ml.) then water (until washings were neutral) and dried in vacuoover phosphorus pentoxide to give the tosylate (120 g., M.P. 147150decomp., [a] +38.6 (c. 1.0, CHCl A mixture of the tosylate (60 g.; 0.124mole) in N,N- dimethylformamide (350 ml.) and potassium acetate (92 g.,0.94 mole) in water (35 ml.) was stirred at for 4 hours. The brownsolution was cooled and most of the N,N-dimethylformamide removed byevaporation at 50 and 4 mm. to give a brown solid mass. Another run withtosylate (58 g., 0.12 mole), potassium acetate (90 g., 0.91 mole),N,N-dimethylformamide (350 ml.) and water (35 ml.) was carried out asdescribed above. The combined residues were shaken with chloroform, thechloroform layer separated and washed with more water (ca. 200 ml.). Thecombined aqueous fractions were extracted with chloroform (3 X 100 ml.)and the combined chloroform extracts (indluding those above) were washedwith water (3 x 100 ml.) and dried over magnesium sulphate. Thechloroform was removed in vacuo and residual N,N dimethylformamide wasevaporated at 50 and 4 mm. to give the crude 3a-acetate (92 g.) as abrown solid.

A solution of the crude acetate (92 g.) in dioxan (1,000 ml.) was mixedwith a solution of potassium hydroxide (45 g., 0.8 mole) in water (500ml.) to give a two-phase system. A homogeneous solution was obtained bythe addition of dioxan (440 ml.) and water (625 ml.). Nitrogen wasbubbled through the solution which was heated at 50 for 2 hours. Theport coloured solution was treated with glacial acetic acid (ca. 40 ml.)to bring the pH to about 7 and two thirds of the solvent was removed bydistillation in vacuo (water pump). Water (ca. 3 l.) was added to theresultant mixtures (which had already begun to crystallise) and theprecipitated solid was filtered off, washed with water and dried overphosphorus pentoxide to give the crude 30t-alCOhOl (73.9 g.).

A solution of the above crude alcohol (73.8 g.) in tetrahydrofuran (1900ml., freshly distilled) with 5% pall'adised charcoal (25 g.) washydrogenated at atmospheric pressure and room temperature. After 1%hours hydrogen uptake (ca. 71) ceased. The catalyst was filtered off ona kieselguhr pad and the filtrate was concentrated in vacuo to acrystalline mass (71.3 g.).

This material was dissolved in chloroform and filtered through a columnof grade III neutral alumina (150 g.) which was washed with chloroform(total ca. 400 ml.). The chloroform was removed in vacuo and the residuetaken up in acetone (200 ml.). Acetone (100 ml.) was boiled off and thesolution was diluted (with stirring with petroleum (ca. 2 1.). Theprecipitated solid was collected by filtration, washed with petroleumand dried to give the crude alcohol (47.1 g.) M.P. 161-163. Retreatmentwith acetone-petroleum gave material (38.3 g.) M.P. 165- 168 and asecond crop (3.0 g.) M.P. 167-168 as fine needles. A further treatmentof these combined crops afforded purer material (33.8 g. plus a 5.3 g.second crop) M.P. 168169 (97.2% pure on G.L.C., 45.5% yield based on3/8-alcohol). A final treatment gave 3a-hydroxy-5apregnane-11,20-dione(35.1 g., 40.8% yield based on 3,9- alcohol; 46.7% yield based on3B-acetate) as needles M.P. 16'8-170 [a] +-1'11.6 (c. 1.0, CHCl (Found:C, 76.15; H, 9.6; Calc. for C H O C, 75.9; H, 9.7%). Purity by G.L.C.analysis was 97.7%.

Chromatography of the combined residues on grade III neutral aluminagave a further amount (9.3 g.) of Bu-hydrOXy-Sa-pregnane-1 1,20-dione.

EXAMPLES 6-16 The table records gas-liquid chromatographic analysis ofmixtures of 3a-acetoxy (and/or formyloxy)-a-pregn- 16-ene-1 1, 20-dione,3u-hydroxy-5a-pregn-l6-ene-l 1,23 dione and5a-pregn-2,16-diene-11,20-dione formed by solvolysis of3B-toluene-p-sulphonyloxy-ia-pregn-l6-ene- 11,20-dione.

12' magnesium sulphate, and concentrated in vacuo to give a whitecrystalline mass. Four recrystallisations from acetone-petroleum (B.P.-60) gave 21-acetoxy-3u-hydroxy-Su-pregnane-l1,20-dione as fine needles(4.22 g., 54%) M.P. 172-173, [u] +104 (c. 1.0. CHCI (Found: C, 70.8; H,8.9; C H 4O requires C, 70.8; H, 8.8%).

EXAMPLE 19 2l-acetoxy-3a-hydroxy-5a-pregnane-11,20-dione via atrimethylsilyl derivative Trimethylchlorosilane (0.33 ml., 1.5 equiv.)and pyridine (0.485 ml., 2 equiv.) were added to a solution of3a-hydroxy-5a-pregnane-11,20-dione (1.0 g.), in methylene chloride (16.6ml.) at room temperature. After 16 hours at 0 more trimethylchlorosilane(0.33 ml. 1 equiv.) was added and after a further 1 /2 hours the solventwas removed in vacuo. Ether was added to the residue and the White solidwas filtered 01f. The ether was removed in vacuo and the residuetriturated with a little petroleum (B.P. 40-60) to give3ot-trirnethyl-siloxy-5upregnane-11,20-dione (0.2 g.), M.P. 9596. Thiscompound was also prepared by the use of hexamethyldisilazane andtrimethylchlorosilane in tetrahydrofuran.

The 21-acetoxylation was carried out as described in Example 18, onecrystallisation from acetone-petroleum (B.P. 4060) alforded the21-acetate, M.P. 170l73.

DISPLACEMENT REACTIONS ON3B-TOLUENE-p-SULPHONYLOXY-51-PREGN-16-ENE-11,20DIONE Percent of- Tem-Weight of ther pera- 3wCH C0 Total tosylate reactants Time ture, pluspercent Per- (grams) Solvent volume (mL) (grams) (hours) C. 3a-HCO 3a-OH3a-CH C0 cent,A 0.56 D.M.F., 10 Lizo a ,0-64-.- 3 115 43.7 12.8 56. 539. 5 1. 9 D.M.F., 75; 11 0.. 46% then 5 80, 90, 64. 5 14. 1 78. 6 21. 4

then 18%. 100 0.58 D.M.F., 15; H20, 0. 11 52. 3 20. 5 72. 8 27. 2 0.532D.M.F., 15; E 0, 1. 27.3 39.4 66. 7 19.7 0. 532 D.M.F., 15; 10, 53.6 21.9 75. 5 24. 5 0.532 D.M.F., 15;HO, 1.2 53.7 17. 8 71. 5 28. 5 0.532D.M.F., 15; H2O, 1.2- "J10"... 27%. 100 32.0 37.6 69. 6 28.0 0.532D.M.F., 15; H2O, 1.2. KOAc, 0.66..- 46% th 80,90 53. 4 21. 5 74. 9 25. 160.0 D.M.F., 35 H20, 35". K AC, 9 -0..- 4 115 59. 7 14. 7 74. 4 23. 7 0.532 D.M.F., 15; E20, 1.2..- KOAO, 0-66- 3 115 50. 3 10.3 69. 6 27.6 0.486 HCOZH, 20; E20, .5 KH a, 0. 5 4 100 44.1 1. 9 46. 0 49.6

EXAMPLE 17 EXAMPLE 20 3u-hydroxy-5a-pregn-16-ene-11,20-dione5'a-pregna-2,16-diene-l1,20-dione (700 mg., 2.2 mmole) was dissolved intetrahydrofuran (10 ml.), and water (4 ml.) was added. Mercuric acetate(700 mg., 2.2 mmole) was added to the stirred solution to give animmediate yellow precipitate. The reaction mixture was stirred at roomtemperature for 24 hours, and then 3 M sodium hydroxide solution (3 ml.)and 0.5 M sodium borohydride in 3 M sodium hydroxide solution (about 3ml.) were added until all the yellow complex had been reduced. Thetetrahydrofuran layer was separated, washed with sodium chloridesolution, and then diluted with ether (100 ml.). The organic solutionwas washed Well with water, dried, and evaporated to give a white oilysolid, which was purified by preparative thin-layer chromatography andrecrystallisation from ethyl acetate/petroleum to give3u-hydroxy-5u-pregn-16-ene-11,20-dione (160 mg., 22%) as colourlesscrystals, M.P. and mixed M.P. with an authentic sample, 234237, [a]+86.5 (c. 1.1, CHCl EXAMPLE 1821-acetoxy-3ot-hydroxy-5ot-pregnane-11,20-dione Boron trifluorideetherate (37.9 ml.) was added to a stirred solution of 3a-hydroxy-5a-pregnane-11,20-dione- (6.64 g., 20 mmole) and leadtetraacetate (10.1 g., 22 mmole) in dry benzene (280 ml.) and methanol(15.1 ml.) at room temperature. After 2 hours the mixture was pouredinto water (2 l.) and extracted with ether (1 1.). The combined etherextracts were washed successively with sodium bicarbonate solution andwater, dried over 2,2,2-trichloroethylchloroformate (1.1 ml.) was addedto a cooled, stirred, solution of 3a-hydroxy-5u-pregnane- 11,20-dione inpyridine (0.74 ml.) and dioxan (5.6 ml.). After 10 minutes dilutehydrochloric acid (14.5 ml.) was added and the solution was heated atfor 30 minutes. The resulting oil crystallised on cooling to give3a-(2,2,2- trichloroethoxycarboyloxy) 50c pregnane 11,20-dione (1.45g.), M.P. -131", [121 -|-56 (c. 1.0, CHC13). (Found: C, 56.0; H, 6.5;Cl, 21.1; C H Cl O requires C, 55.6; H, 6.6; Cl, 20.7%

21-acetoxylation of this compound (0.5 g.), using the proceduredescribed in Example 18 gave a syrupy residue that crystallised on theaddition of ether and petroleum. Recrystallisation from ether-petroleum(B.P. 40-60) gave 21-acetoxy-3ot-(2,2,2 trichloroethoxycarbonyloxy)-Sa-pregnane-ILZO-dione (0.325 g.), M.P. 143146 (decomp.), [04] +75.2 (c.1.0, CHCl (Found: C, 55.1; H, 6.0; Cl, 18.6. C25H35C13O7 requires C,54.3; H, 6.4; Cl, 19.2%

Removal of the 2,2,2-trichloroethoxycarbonyl group was effected bystirring a solution of the steroid (0.10 g.), in ethanol (2 ml.) andglacial acetic acid (2.5 ml.) for 5 hours with addition of zinc dust(0.3 g.) during this period. The solids were filtered 01f, washed withethanol and the combined filtrate was reduced in volume in vacuo.Addition of water to the residual solution gave 21-acetoxy-3a-hydroxy50c pregnane-ll,20-di0ne (0.036 g.), M.P. 168l72.

13 EXAMPLE 21 21-acetoxy-5 a-pregnan-3a-ol-1 1,20-dione Dried yeast(Saccharomyces cerevisiae 50 g.) was stirred at room temperature withtap water (2 1.) containing sucrose (200 g.) and di-ammonium hydrogenphosphate (4 g.). After two hours 21-acetoxy-5a-pregmane-3,11,20-trione(1 g.) (Mancara et al., J. Amer. Chem. Soc., 1955, 77 5669) in ethanol(100 ml.) was added. The fermentation was stirred at room temperaturefor a further 24- hours, the yeast was removed by filtration, partiallydried at the pump and extracted with hot chloroform (4 250 ml.). Theextracts were shaken with the filtrate, washed with water, dried (Na SOand evaporated to a brown gum (634 mg). This was subjected topreparative thin-layer chromatography which yielded two pure substances.Of these the less polar was the starting material21-acetoxy-5u-pregnane-3,11,20-trione, (68 mg); which was recrystallisedfrom acetone/hexane as colourless needles, M.P. 169-172; [4x1 127.S, (c.0.87 CHCl Gas-liquid chromatography confirmed that this was identicalwith the starting material.

The more polar substance (70 mg.) was recrystallised from acetone/hexaneto afford 21-acetoxy-5u-pregnan- 3a-ol-11,20-dione (57 mg.) ascolourless needles, M.P. 179-182"; [ab +107", (c. 1.09 CHCl Gas-liquidchromatography confirmed the identity of this material.

EXAMPLE 22 21-acetoxy-5a-pregnan-3u-ol-11,20-dione (a) 21-bromoSa-pregnan-3a-I1,20-dione.5a-pregnan-3tx-o1-11,20-dione (1 g., 3 mmoles)in stirred methanol (7 ml.) at 30 was treated with acetyl chloride (1drop). After two minutes bromine (0.19 ml., 3.52 mmoles) in methanol(4.5 ml.) was added dropwise, the solution being allowed to decolourisebetween the addition of each drop. The resulting clear solution waspoured into chloroform (100 ml.), washed with water (3X 50 ml.), dried(Na SO4) and evaporated to a white froth (1.40 g.). Preparativethin-layer chromatography afforded21-bromo-5ot-pregnan-3a-ol-l1,20-dione (715 mg.) which crystallised fromchloroform/ether as clusters of colourless needles, M.P. 160l63; [a1+l09 (c. 0.82, CHCl (Found: C, 61.0; H, 7.9; Br, 19.7; C H BrO requiresC, 61.3; H, 7.61; Br, 19.45%).

A second, unstable substance (415 mg.) isolated from this reaction wasexamined spectroscopically and identified as17a-bromo-5wpregnan-3ot-ol-11,20-dione.

(b) 21-acetoxy 50c pregnan-3a-ol11,20-dione.5otpregnan-3a-ol-l1,20-dione(1 g. 3 mmoles) was brominated in the manner described above. The totalproduct from the reaction (1.38 g.) was refluxed with stirring with dryacetone (25 ml.) and anhydrous potassium acetate (2 g.). After fourhours the reaction mixture was poured into chloroform (100 ml.), washedwith water (3x 50 ml.), dried (Na SO and evaporated to a white froth(1.19 g.). Preparative thin-layer chromatography afforded21-acetoxy-5a-pregnan-3a-ol-11,20-dione (634 mg.) as a white froth whichon crystallisation from acetone/ether gave colourless needles, M.P.177-180; [111 +l02.5, (c. 0.95 CHCI Gas-liquid chromatography confirmedthe identity of this material.

EXAMPLE 23 21-acetoxy-5 a-pregnan-3 ot-l 1,20-dione5a-pregnan-3a-ol-l1,20-dione (5 g., 15.0 mmoles) in stirred methanol (35ml.) at was treated with drops of a 50% w./v. solution of hydrogenbromide in glacial acetic acid. After several minutes, bromine (0.95ml., 2.815 g. 17.6 mmoles) in methanol (22.5 ml.) was added portionwiseover ca. minutes. The solution was left stirring until the colourdisappeared and then poured into water. The product was isolated byfiltration, washed with water and dried in vacuo at 40 for five hours.Final drying was over phosphorus pentoxide in a desiccator (overnight).Yield: approximately quantitative.

The crude bromo-intermediate was dissolved in acetone ml., dried overpotassium carbonate) and refluxed in the presence of potassium acetate(10 g.) for four hours.

The reaction mixture was poured into water (1 l.) and extracted withether (300 ml. and 250 ml.). The ether was dried over anhydrousmagnesium sulphate, filtered and evaporated under reduced pressure.

The residual foam was recrystallised from acetone-petroleum ether togive 2l-acetoxy-5a-pregnan-3a-ol-l1,20- dione as white needles. Yield,63%; M.P. 173175; [0:1 109', (c. 1.0, CHClg).

EXAMPLE 24 21-propionyloxy-5a-pregnan-3 0L-Ol-11,20-dl01'165ot-pregnan-3 ot-ol-11,20-dione (2 g., 6 mmoles) in methanol (14 ml.) at30 was treated with acetyl chloride (2 drops). The solution was stirredfor two minutes and was treated dropwise with bromine (0.38 ml., 7mmoles) in methanol (9 ml.) in the manner described above. The reactionmixture was poured into stirred water (250 ml.). The product wasisolated, washed and dried at the pump and redried over phosphoruspentoxide in vacuo to afford a white powder (2.49 g.). This was refluxedwith propionic acid (11.1 ml., 0.15 mmoles) and triethylamine (7.25 ml.,0.1 mole) in dry acetone (60 ml.). After five hours the solution waspoured into chloroform (300 ml.), washed with aqueous potassium hydrogencarbonate (2X 200 ml.) and with water (2 200 ml.) dried (Na SO andevaporated to a white froth (2.59 g.). Preparative thin-layerchromatography afforded material which on recrystallisation fromchloroform/ether/ hexane gave21-propionyloxy-Swpregnan-Su-ol-11,20-dione (589 mg.) as colurlessirregular prisms, M.P. 159-170"; [ab +99, (c. 0.92 CHCl (Found: C, 70.5;H, 8.9. C H O JAH O requires C, 70.45; H, 9.0%).

EXAMPLE 25 21-isobutyryloxy-5a-pregnan-3u-ol-l1,20-dione5a-pregnan-3a-ol-l1,20-dione (2 g., 6 mmoles) in methanol (14 ml.) at 30was treated with acetyl chloride (2 drops). The solution was stirred fortwo minutes before being treated dropwise with bromine (0.38 ml., 7mmoles) in methanol (9 ml.) in the manner described above. When thereaction was complete the solution was poured into stirred water (25 0ml.). The product was isolated, washed and dried at the pump. A finaldrying over phosphorus pentoxide in vacuo afforded a white powder (2.5g.). This was refluxed with isobutyric acid (14.1 ml., 0.15 moles) andtriethylamine (7.25 ml., 0.10 mole) in dry acetone (60 ml.). After fourhours the solution was poured into chloroform (300 ml.), washed withaqueous potassium hydrogen carbonate (2X 200 ml.), with water (2 200ml.), dried (Na SO and evaporated to a white froth (2.54 g.).Preparative thin-layer chromatography afiorded 21isobutyryloxy-5a-pregnan-3a-ol-l1,20-dione (1.45 g.) as a white froth,[a1 +93.5, (c. 0.66 CHCl (Found: C, 70.4; H, 8.8. C H O /2H O requiresC, 70.25; H, 9.2%).

EXAMPLE 26 21-benzoyloxy-5a-pregnan-3 a-ol-l 1,20-dione5oc-pregnan-3a-ol-l1,20-dione (2 g., 6 mmoles) in methanol 14 ml.) wastreated with acetyl chloride (2 drops) and bromine (0.38 ml., 7 mmoles)as described above. Isolation of the crude bromination product in theusual manner afforded a white powder (2.498 g.). This was refluxed withbenzoic acid (18.3 g., 0.15 moles) with triethylamine (7.25 ml., 0.1moles) in dry acetone (100 ml.). After five hours the solution waspoured into chloroform (500 ml.), washed with aqueous sodium hydrogencarbonate (2X 200 ml.) and with water (3X 200 ml.), dried 15 (Na- SO andevaporated to a white froth. Preparative thin-layer chromatographyafforded 21-benzoxyloxy-5apregnan-3ot-ol-11,20-dione (1.27 g.) as awhite froth, [01.1 +140, (c. 0.65, CHCl (Found: C, 72.6; H, 7.7. C H O/2H-, O requires C, 72.85; H, 8.05).

EXAMPLE 27 21isobutyryloxy-Soc-pregnan-3 u-ol-l 1,20-dione 3-nitrate (i)Fuming nitric acid (1.3 ml.) was added to stirred acetic anhydride (5ml.). The solution was cooled to 5 and21-acetoXy-5u-pregnan-3a-ol-11,20-dione (500 mg. 1.28 mmoles) inchloroform (2.5 ml.) was added.

The resulting solution was stirred at 5 for an hour before being pouredinto dilute aqueous sodium hydroxide (100 ml.), extracted withchloroform (2 75 ml.) and the extract washed with aqueous sodiumbicarbonate (50 ml.), with water (3X 50 ml.), dried (Na SO andevaporated to a gum (527 mg.). Crystallisation from ether/ acetoneafforded 2l-acetoxy-5a-pregnan-3a-ol-11,20-dione 3-nitrate (386 mg.) ascolourless needles, M.P. 147- 149; [ab 103 (0. 1.06 CHCl (Found: C,63.6; H, 7.5; N, 3.1. C H NO requires C, 63.45; H, 7.65; N, 3.2%).

(ii) 5-u-pregna-3a,2l-diol-11,20-dione 3-nitrate (a)21-acetoxy-5a-pregnan-3a-ol-l1,20-dione 3-nitrate (837 mg., 2 mmoles) inethanol (42 mg.) and tetrahydrofuran (42 ml.) was stirred with 10%aqueous potassium hydrogen carbonate (4 m1.) and 2 N aqueous sodiumhydroxide (2 ml.). After three hours glacial acetic acid (1 ml.) wasadded, the solution was poured into water (400 m1.) and extracted withchloroform (3x 120 ml.). The extracts were washed with water (2 50 ml.),dried (MgSO and evaporated to a froth. Preparative thinlayerchromatography afforded pure material which on recrystallisation fromether/acetone gave 5a-pregna-3u, 21-diol-11,20-dione 3-nitrate (219 mg.)as colourless irregular prisms, M.P. 172-176"; [ah +91 (c. 0.97, CHCl(Found: C, 63.9; H, 7.8; N, 3.5. C H NO requires C, 64.1; H, 7.95; N,3.55%

(b) 21-acetoXy-5a-pregnan-3a-ol-11,20-dione (5 g., 13.3 moles) inchloroform (25 ml.) was added to a stirred solution of fuming nitricacid (13 ml.) in acetic anhydride (50 ml.) at 5. The reaction mixturewas stirred at 5 for an hour, poured into stirred aqueous sodiumhydroxide (1 l.) and extracted with chloroform (2 200 ml.). The extractwas washed with aqueous sodium hydrogen carbonate (100 ml.) and withwater (2 100 ml.), dried (Na SO and evaporated to a white froth. Thiswas dissolved in methanol (500 ml.), the solution flushed with nitrogenand stirred with 10% aqueous potassium hydrogen carbonate (17.5 ml.) forfour hours. Glacial acetic acid (3 ml.) was added, the solutionevaporated to small bulk, poured into water (1 l.) and extracted withchloroform (2 200 ml.). The extract was washed with water (2 100 ml.),dried (Na SO and evaporated to a white froth. Crystallisation fromacetone/ ether gave 5a-pregna-3u,2l-diol-11,20-dione 3-nitrate (4.31 g.)as colourless irregular prisms, M.P. 174-181".

(iii) 21isobutyryloxy-5ot-pregnan-3u-ol-l1,20=dione 3-nitrate5a-pregna-3a,21-diol-11,20-dione 3-nitrate (2.0 g., 5.08 mmoles) in drypyridine ml.) was treated with isobutyryl chloride (2 ml.). The pinksolution was allowed to stand at room temperature for 20 hours beforebeing poured into stirred water and acidized with dilute hydrochloricacid. The product was isolated, washed thoroughly with water and driedat the pump. Recrystallisation from acetone/hexane afforded21isobutyryloxy-5a-pregnan-3uol-l1,20-dione 3-nitrate (1.01 g.) ascolourless needles, M.P. 167169; [01.] +97.5 (c. 1.48, CHCl (Found: C,64.5; H, 7.8; N, 3.2. C H NO requires C, 64.8; H, 8.05; N, 3.0%).

1 6 (iv) 21isobutyryloxy-Sm-pregnan-3 a-ol-1l,20-dione 2lisobutyryloxy-Sa-pregnan-3 a-ol- 1 1,20-dione 3-nitrate (1.35 g., 2.91mmoles) in glacial acetic acid (35 ml.) was stirred for an hour at roomtemperature with zinc powder (4 g.). The zinc was removed, washed withhot chloroform (200 ml.) and the combined washings and filtrates werewashed with water (4x100 ml.) dried (Na SO and evaporated to a whitefroth. Preparative thin-layer chromatography afforded21isobutyryloxy-Sa-pregnan- 30L-Ol-11,20-di0l18 as a white froth, [aJ+97.5 (c. 1.05, CHCl Analytical thin-layer chromatography and gasliquidchromatography indicated that this material was identical With a sampleprepared by the method described in Example 31.

EXAMPLE 28 3u-hydroxy-5u-pregn-16-ene-1 1,20-dione and Soc-pregna-2,16-diene-1 1,20-dione A stirred mixture of3,8-toluene-p-sulphonyloxy-5mpregn16-ene-l1,20-dione (627 g.), potassiumacetate (918 g.), dimethylformamide (4.25 l.) and water (425 ml.) washeated on the steam bath for 4 hrs. Most of the dimethylformamide wasremoved under reduced pressure and water was added to the residue withstirring. The acid (420 g.) was collected, washed and dried at 40 invacuo. This material in peroxide free dioxan (7 l.) was flushed withnitrogen and a solution of potassium hydroxide (200 g.) in water (2 l.)was added. The mixture was stirred and heated at 50-60" for 7 hrs. andthen left at room temperature overnight. Acetic acid (50 ml.) was addedand the precipitated solid filtered olf, washed with water and dried.The dry solid was heated under reflux with benzene (2 l.) for 2 hrs. themixture cooled and filtered to give30chydroxy-5ot-pregn-16-ene-11,20-dione (215 g.).

Benzene extracts from several similar experiments were combined andevaporated. The crude solid was redissolved in hot benzene and petroleum(B.P. 60-80) was added until the solution was just clear at ambienttemperature (an insoluble residue was removed by filtration at thisstage). The clear solution was passed down an alumina column (Woelmalumina, activity grade I) previously prepared with benzene-petroleum(B.P. 6080) in the ratio 6:4. Elution with the same solvent gave thediene. Elution was terminated when thin layer chromatography indicatedthe presence of slower running components in the eluate. The combinedfractions were evaporated and twice crystallised from benzene-petroleum(B.P. 6080) to give 5u-pregna-2,16-diene-l1,20-dione as colourlesscrystals, MJP. 176-177, [ab +159 (0. 1.0 in CHClg), A (EtOH) 233.5 nm.(e 9150); (Found: C, 80.5; H, 8.9; C21H2g02 requires C, H,

EXAMPLE 29 5 u-pregna-3a,21-diol-1 1,20-dione 21-hemisuccinate5ot-pregna-3u,2l-diol-11,20-dione 3-nitrate ZI-hemisuccinate (1.03 g.,2.09 mmoles) and zinc powder (3 g.) were stirred with acetic acid (25ml.) for minutes. The zinc was removed, washed with chloroform ml.), 2 Naqueous hydrochloric acid (200 ml.) and chloroform (100 ml.). Thecombined filtrates were equilibrated and the organic phase was washedwith Water (3 X100 ml.), dried Na SO and evaporated to a white froth.Crystallisation from benzene/hexane/chloroform gave 5oa-pregna-3a,21-diol-11,20-dione zl-hemisuccinate (800 mg.) as colourless needles, M.P.166-l68; [ab +90.5 (c. 1.32 in CI-ICl (Found: C, 66.9; H, 8.0. C H O,requires C, 66.95; H, 8.1%.

The starting material for this preparation was obtained from5a-pregna-3a,21-diol-11,20-dione 3-nitrate (see Example 33ii) asdescribed below:

5ot-pregna-3a,21-diol-11,20-dione 3-nitrate (2 g., 5.08 mmoles) andsuccinic anhydride (2 g. 20 mmoles) in dry pyridine (20 ml.) wereallowed to stand at room temperature for 18 hours. The solution waspoured into stirred water (300 ml.) which was then acidized. After twohours, the precipitate was collected, washed well with water and dried.Recrystallisation from benzene/hexane gave5wpregna-3a,21-diol-11,20-dione 3-nitrate 21-hemisuccinate (1.51 g.) ascolourless micro-needles; MP. 160- 163"; [th, +89.5 (c. 1.14, CHCl(Found: C, 60.8; H, 7.3; N, 2.6. C H NO requires C, 60.8; H, 7.15; N,2.85%).

EXAMPLE 30 0.03 g. of steroid I, finely divided in an air-attrition milland having an approximate mean particle diameter of Sn, was mixed with 2g. of Cremophor EL by mechanical agitation and heated to 70 in anatmosphere of nitrogen. Heating and agitation was continued until thesteroid had dissolved. The solution was diluted with sterile distilledwater containing 0.025 g. of sodium chloride to gain a final volume of10 ml., and stirring continued until the solution was homogeneous.

EXAMPLE 3 1 0.035 g. of steroid I reduced in size as in Example 30 mixedwith 0.035 g. of steroid II (R=CH CO) reduced in size as for steroid I.The steroids are added to 10 ml. of a 20% w./v. solution of Chemophor ELcontaining 0.025 g. of sodium chloride and the mixture mechanicallystirred for at least 24 hours. Any traces of insoluble residue arefiltered off using a sintered glass filter.

EXAMPLE 32 13.5 g. of ground steroid I and 4.5 g. of steroid II (R=CHCO-) are mixed by mechanical agitation with 300.0 g. of Cremophor EI at70 C. in an atmosphere of nitrogen. Heating and agitation are continueduntil the steroids have completely dissolved. The mixture is dilutedwith sterile distilled water containing 3.75 g. of sodium chloride togive a final volume of 1500 ml. Stirring is continued until the solutionis homogeneous.

EXAMPLE 33 0.045 g. of steroid I and 0.015 g. of steroid II (R=CH CO)are dissolved in 2 ml. of acetone at 20 C. The resultant solution isadded to 1 g. of Cremophor EL at 20 C. and stirred until homogeneous.The acetone is removed by a vigorous stream of nitrogen. The solution isdiluted with sterile distilled water containing 0.0125 g. of sodiumchloride to give a final volume of ml. Similar solutions were preparedusing chloroform or methylene chloride in place of acetone.

EXAMPLE 34 0.09 g. of ground steroid I are mixed with 0.03 g. of groundsteroid II (R=CH CO) and this added to 2.0 g. of Tween 80. As in Example33, the mixture is mechanically agitated at 70 C. in a stream ofnitrogen until the steroids have dissolved. The mixture is diluted withsterile distilled water containing 0.025 g. of sodium chloride to give afinal volume of ml.

EXAMPLE 35 0.045 g. steroid I and 0.015 g. of steroid II (R: (CH CHCO)are formulated as in Example 33. Similar solutions are obtained usingsteroid II wherein R=C H CO, H0 OCH CH CO or C H -C0.

EXAMPLE 36 0.045 g. of steroid I and 0.015 g. of steroid II (R=CH CO)are dissolved in 2 ml. of acetone at C. The resulting solution is addedto 1 g. of Tween 40 at 20 C. and stirred until homogeneous. The acetoneis removed by a vigorous stream of nitrogen. The solution is dilutedwith sterile distilled water containing 0.025 g. of sodium chloride togive a final volume of 10 ml.

EXAMPLE 37 0.036 g. of ground steroid I are mixed with 0.012 g. ofground steroid II (R=CH CO). This is added to 0.8

18 g. of Tween 60. The mixture is mechanically agitated at 70 C. in astream of nitrogen until the steroids are dissolved. The mixture isdiluted with sterile distilled water containing 0.025 g. of sodiumchloride to give a final volume of 10 ml.

EXAMPLE 38 16 mg. of steroid I and 4 mg. of steroid II (R=CH CO) weredissolved in 0.2 ml. of acetone, and 0.2 ml. of Cremophor EL was mixedwith this solution. The acetone was removed by passing a stream ofnitrogen through the mixture. The resulting solution was mixed with 0.8ml. of sterile water.

EXAMPLE 39 0.135 grams Steroid I and 0.045 gram of Steroid II, (R=CHCO), are dissolved in 2 mls. of acetone at 20 C. The resulting solutionis added to 3 grams of Cremophor EL at 20 C. and stirred untilhomogeneous. The acetone is removed by a vigorous stream of nitrogen.The solution is diluted with sterile distilled water containing 0.025grams of sodium chloride to give a final volume of 10 mls.

EXAMPLE 40 0.045 grams of ground Steroid I are mixed with 0.012 grams ofground Steroid II (R=CH CO), and the mixture added to 1 gram ofCremophor EL. The mixture is mechanically agitated at 70 C. in a streamof nitrogen until the steroids have dissolved. The mixture is dilutedwith sterile distilled water containing 0.025 grams of sodium chlorideto give a final volume of 10 mls.

We claim:

1. A compound of the general formula CH1 0 R wherein R is selected fromthe group consisting of a lower alkanoyl group, a hemisuccinoyl groupand a benzoyl group.

2. A compound as claimed in claim 1 wherein R is an alkanoyl groupcontaining from 2 to 4 carbon atoms.

3. A compound as claimed in claim 1 which is 21- acetoxy-3a-hydroxy-5a-pregnanel 1,20-dione.

4. A compound as claimed in claim 1 which is3a-hydroxy-2l-propionyloxy-Sa-pregnane-1 1,20-dione.

5. A compound as claimed in claim 1 which is 21-isobutyryloxy-3u-hydroxy-5ot-pregnane-l 1,20-dione.

6. A compound as claimed in claim 1 which 3a-hydroxy-5a-pregnane-l1,20-dione 2 l-hemisuccinate.

7. A compound as claimed in claim 1 which is 21- benzoyloxy-3a-hydroxy-5u-pregnane-1 1,20-dione.

OTHER REFERENCES J.A.C.S., vol. (1958), p. 250, by Ringold et al.

ELBERT L, ROBERTS, Examiner US. Cl. X.R.

2 33 EJ'LKHTED STATES PATENT 0mm; CER'FEFNIA'FE )F {IQMQECTWN Patent No3, 1 m a Octobez: 2, 1973 Inventoris) smwmm DAVIS, BEFEEZK RGGER PEARCEet a1 It. is certified that error appears in the above-identified patentand that said Letters Patent are hereby corrected as shown below:

(Jolumn 1,? limos 3-11'3, inclusive, should read Bsnjamin Davis, {:2font St, L Qter Desek Rogax': Pearce, mfaagckmollg rfiiall Galbrsi'izhWail, London Gordon Ian GET'BQ'QZPSY, Chulfont St, matsr, and; GowffionHanlsy Flifiipps, Wembls'y', England, assignors to Galxo LsboxatorissLimitsd, Grssnfosd Middlssex, England Column 18, ths formula set; forthat: lines 35-47 should appear as follows:

E -I QR Signed and sealed this 12th day of November 1974.

(SEAL) Aiitest:

McCOY M. GIBSON JR. C. MARSHALL DANN Attesting Officer Commissioner ofPatents

